Inflammatory reaction is one of the body's responses deeply involved in the symptoms of many diseases, and is now an important subject of study in understanding their pathology and/or in developing therapeutic strategies against these diseases. For this reason, it is indispensable to develop a technique which allows detection of the actual conditions of inflammation.
A typical inflammatory reaction has been elucidated to occur through the following mechanism. Namely, once a source of infection (e.g., bacteria, viruses) has entered the body, the source of infection will be detected by cell surface receptors, followed by induction of cytokine secretion. The secreted cytokines will serve as guides to cause immunocytes (e.g., macrophages) to migrate to the infection site, whereby the source of infection will be eliminated. The increased activity of these immunocytes during elimination will result in flare, fervescence, pain and swelling which are characteristic of inflammation.
As a cytokine which is significantly involved in this inflammatory reaction and also receives attention as an inflammatory marker, interleukin-1 beta (IL-1β) has been known. IL-1β is not substantially secreted in the absence of inflammatory stimulation, but is known to be produced and secreted at a very high level in each tissue upon inflammatory stimulation (FIG. 1).
IL-1β has been found to be strictly regulated by the following characteristic two-stage control. The gene expression of IL-1β is activated by transcription factor NF-κB induced during inflammatory reaction, and the activated IL-1β gene expression in turn promotes the production of precursor proIL-1β. Then, proIL-1β will be cleaved by caspase activated in inflammasomes and converted into secretable mature IL-1β (FIG. 2).
Some reports have been issued about the monitoring of IL-1β gene expression in which luciferase or red fluorescent protein is used as a reporter molecule (Non-patent Documents 1 and 2). In these reports, it is shown that transgenic mice carrying a reporter molecule are prepared and in this inflammation model, reporter signals can be detected and also can be used for in vivo imaging analysis. However, this method relies only on transcriptional regulation, which is one factor contributing to a cascade of inflammatory reactions, and hence this method is insufficient to monitor physiological inflammatory reactions.
On the other hand, a reporter system regulated by inflammasomes has also been reported (Non-patent Document 3). In this report, a reporter molecule is design such that it is in an inactive state due to aggregation in the absence of inflammation, but it will be converted into a monomer form to exert its activity when inflammasomes become functional upon inflammatory stimulation. However, this system also relies only on inflammasomes and is therefore insufficient in sensitivity. Moreover, this system has not been verified as to whether it is functional in living mice.
Other attempts have also been made to induce protein expression by various internal or external stimuli (e.g., oxidative stress, endoplasmic reticulum stress) and to visualize the event taking place (Patent Documents 1 and 2, Non-patent Documents 4 and 5).